5e2j | pdb_00005e2j

Crystal structure of single mutant thermostable endoglucanase (D468A) from Alicyclobacillus acidocaldarius

spinqualitylabelsall models 5e2j, resolution 2.10Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 5e2j is a 2 chain structure with sequence from Alicyclobacillus acidocaldarius subsp. acidocaldarius. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 2.1Å Ligands: CA, MPD, ZN Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function Q9AJS0_ALIAC Publication Abstract from PubMed Structural calcium sites control protein thermostability and activity by stabilizing native folds and changing local conformations. Alicyclobacillus acidocaldarius survives in thermal-acidic conditions and produces an endoglucanase Cel9A (AaCel9A), which contains a calcium-binding site (Ser465-Val470) near the catalytic cleft. By superimposing the Ca2+-free and Ca2+-bounded conformations of the calcium site, we found that Ca2+ induces hydrophobic interactions between the calcium site and its nearby region by driving a conformational change. The hydrophobic interactions at the high B-factor region could be enhanced further by replacing the surrounding polar residues to hydrophobic residues in turn to affect enzyme thermostability and activity. Therefore, the calcium-binding residue Asp468, whose side chain directly ligates Ca2+, along with Asp469 and Asp471 of AaCel9A, were separately replaced by alanine and valine. Mutants D468A and D468V showed increased activity compared with those of wild type at 0 mM or 10 mM Ca2+; whereas the Asp469 or Asp471 substitutions resulted in decreased activity. The D468A crystal structure revealed that mutation D468A triggered a conformational change similar to that induced by Ca2+ in wild type and developed a hydrophobic interaction network between the calcium site and the neighboring hydrophobic region (Ala113-Ala117). Mutations D468V and D468A increased 4.5 degrees C and 5.9 degrees C in melting temperature, respectively; and enzyme half-life at 75 degrees C increased approximately 13 times. Structural comparisons between AaCel9A and other endoglucanases of GH9 family suggested that the stability of the regions corresponding to the AaCel9A calcium site plays an important role in GH9 endoglucanase catalysis at high temperature. Polarity Alteration of Calcium Site Induces a Hydrophobic Interaction Network and Enhances Cel9A Endoglucanase Thermostability.,Wang HJ, Hsiao YY, Chen YP, Ma TY, Tseng CP Appl Environ Microbiol. 2016 Jan 4. pii: AEM.03326-15. PMID:26729722 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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